Mutagenesis of longer inserts by the ligation of two PCR fragments amplified with a mutation primer

Kato Y; Arakawa N; Masuishi Y; Kawasaki H; Hirano H

Journal of Bioscience and Bioengineering, Vol.107, No.1, 95-97, 2009

DOI10.1016/j.jbiosc.2008.09.003 Export Citation

Mutagenesis of longer inserts by the ligation of two PCR fragments amplified with a mutation primer

Kato Y, Arakawa N, Masuishi Y, Kawasaki H, Hirano H

We report a method for efficient mutagenesis of DNA in large vectors without subcloning. Two segments of the target DNA sequence, one having a mutation introduced via a mutant primer, were amplified by PCR and then the purified fragments were ligated to a vector. The mutation efficiency was nearly 100%. (C) 2008, The Society for Biotechnology. Japan. All rights reserved.

Keywords:Polymerase chain reaction (PCR);Mutagenesis;Yeast;Expression vector;Deletion;Insertion;Substitution